Men and other animals are continually exposed to infections and re-infection by various species and strains of free-living limax amoebae which can be detected in the faeces, nasopharynx and bronchi. In all parts of the world they form part of the environment. Experimentally in animals they induce changes like those of collagen and auto-immune diseases and are characterized by vasculitis, myositis, hepatitis, pyelitis and splenomegaly. They can often be seen in the tissues of animals. Such animals show lymphadenopathy with the appearance like that of human Hodgkin's disease or a state like that of advanced malignant disease. These organism may also be recovered from all the tissues of cases of collagen and auto-immune diseases and from human and many animal tumors and may also occur in the tissues of apparently healthy individuals. They cannot be identified in ordinary sections, but can be demonstrated by immunofluorescent methods.
The definite cause of rheumatoid arthritis is presently unknown. Rheumatoid arthritis is a crippling disease, characterized by the inflammation of several joints of the body, with swelling, pain and stiffness. Rheumatoid arthritis is a disorder that afflicts about fifteen million people in the Western World alone. Successful early treatment may avert the destructive, deforming phase of the disease. Therapy has been directed largely at non-specific suppression of inflammatory and immunologic processes. Aspirin is the cornerstone of therapy for rheumatoid arthritis and can reduce pain in a majority of patients in view of its analgesic action. Widespread interest in rheumatoid arthritis arose when Hench (1949) introduced cortisone in treatment. Chemical compounds which have been commonly used in treating rheumatoid arthritis are corticosteroids, gold salts, anti-malarial drugs, immunosuppressive agents and a whole range of so-called non-steroidal drugs, e.g. indomethacin, phenylacetic acid (Ibuprofen), propionic acid (Naproxen) and D-Penicillamine. Most of these drugs being temporary relief to the arthritic patient but present the danger of side effects and the physician has to balance the potential benefit against the risks. However, arthritis reoccurs following withdrawal of such chemical treatment. For many years rheumatoid arthritis was considered to be an infection (Hollander et al., 1960; Robinson, 1967), but with the advent of the concept of auto-immunity this idea lost favor. Such a view has recently been revived (Lancet, 1970, 2, 303) and is supported by many observations. It is highly likely that the limax amoebae, found in all the collagen and auto-immune diseases, may well be the aetiological agent of these conditions and that anti-protozoal drugs may help by their action on these organisms.
The use of a bis-phenyl (2-halophenyl)-1-imidazolylmethane (e.g. clotrimazole) for the treatment of rheumatoid arthritis is disclosed in my U.S. Pat. No. 4,073,922 issued on Feb. 14, 1978. The use of tinidazole and related compounds is disclosed in my U.S. patent application Ser. No. 813,922, filed July 8, 1977, now U.S. Pat. No. 4,119,723. It has also been suggested to use a nitroimidazole derivative in the treatment of rheumatoid arthritis in the Journal of Tropical Medicine and Hygiene v. 75, p. 64 to 66, Mar. 1972.
Various other anti-protozoal drugs were tried on cases of rheumatoid diseases or of various localized manifestations of this. The substances investigated were 4-aminoquinolines (chloroquine), hydroxychloroquine (plaquenil), amodiaquine (camoquin), copper sulfate and bile salts (dehydrocholine), which are effective in killing the trophozooites of many amoebae in the concentration found in the small intestine. All of these were actually shown experimentally to kill amoebae. In addition, other anti-protozoal drugs were also investigated. They include suramin, pentamidine, dehydroemetine (DHE or mebadin), metronidazole (flagyl), nimorazole (naxogin), phanquone (entobex) and diloxanide (furamide).
The 4-aminoquinolines have been given orally in a dose of 200 and 400 mg. daily, reduced after a month to 200 mg. twice weekly, care being given to examine the eyes at intervals to guard against keratitis or macular changes. Copper salts were administered as 25 mg. of copper sulfate in aqua chloroformi by mouth three times daily. This may produce vomiting and/or diarrhea and the dose has to be decreased to 10 mg. three times daily. Only a small amount of the metal is absorbed, however, and no other side effects are observed even when taken over several months. Bile salts as dehydrocholine were given in a dose of 500-1000 mg. three times a day by mouth. They may produce mild colic. Pentamidine was at first given by intramuscular injection into the buttock in doses of 200 mg. daily for ten days. The course was repeated twice with intervals of seven days between. This substance is liable to produce local necrosis or abscess formation. Pentamidine can be given by mouth, but the absorption is uncertain. Capsules containing 200 mg. were especially made and a dose of 200 mg. twice daily to 400 mg. three times daily by mouth were tried in various combinations. Suramin was given by intravenous injection of 500 mg. in 10 ml. of water and after this every four hours 1 g. was injected until 10 g. had been given. The course was repeated once after four months. Dehydro-emetine (DHE) was given by intramuscular injection in doses of 60 mg. daily for ten days and repeated after seven days, or 60 mg. three times daily by mouth for 7-10 days, repeated after an interval of ten days. Before commencing treatment E.C.G.'s were taken and repeated before each successive injection. Metronidazole (flagyl) was given in doses of 400 to 600 mg. three times daily by mouth and nimorazole (naxogin) in doses of 75 mg. three times daily. Phanquone (entobex) was given in doses of 100 mg. twice daily by mouth for seven days, repeated at intervals of a week, Diloxanide (furamide) was given in doses of 500 mg. three times daily for ten days and repeated once.
The various substances tested above were tried on cases of rheumatoid arthritis of varying severity, systemic lupus erhthematosus, dermatomyositis and other manifestations of collagen and auto-immune disease and observations made on the clinical condition, oedema, morning stiffness, E.S.R., plasma proteins, RF, ANF and organ-specific antibodies in the serum. No attempt at a double-blind trial was made as its became obvious, fairly early or even the day after commencing treatment, whether beneficial results were obtained and furthermore, symptomatic improvement is associated with improvement or disappearance of the abnormal blood changes, indicating that the drug was effective and improvement not due to suggestion. No beneficial effect was obtained from flagyl, naxogin, entobex, suramine or furamide in the doses used. However, Abd-Rabbo et al. (1972), using a derivative of nitro-imidazole BT 985 Merck A.G., which is active against amoebae, giardia and trichomonas, obtained beneficial effects in one case of systemic lupus erythematosus and nine of ten cases of rheumatoid disease. The drug was given in doses of 250 mg. daily for 14-39 days. In the follow up period of 3-6 months no treatment was given and it was noted that the pain recurred, yet not to the same degree as before the treatment.
It has now been discovered that certain substituted imidazole compounds which have anti-protozoal activity are effective for treating rheumatoid arthritis and related collagen and auto-immune (rheumatoid) diseases.
The method of determining the anti-protozoal activity of drugs on limax amoebae was described by Fulton, C. Methods in Cell Biology (edited by D. M. Prescott), p. 341, New York, 1070, and followed by Jamieson and Anderson in Lancet, 1974, 1, 261. All experiments were performed using 5-day old 5 ml. cultures of amoebae in the axenic medium "A" of Fulton. A standard inoculum of 100 c.mm. of differing concentrations of amoebae was added to each 1 ml. tube containing the dilutions of the compound to be tested (dissolved initially in dimethyl sulfoxide) or other drugs in the axenic medium. The tubes were incubated for 5 days at 37.degree. C. and the final concentrations per c.mm. was compared with the initial count to determine percentage kill.